FIELD OF THE INVENTION the Invention
This invention relates to an immunochromatography using dyed latex particles of a synthetic high polymer as fine particles to which an antigen or antibody is to be bound.
As a simple immunoassay for detecting a substance to be detected which consists of a specific antigen or antibody by use of a specific reaction between antigen and antibody, there has heretofore been used an agglutination method which comprises binding, by immunoreaction, a substance to be detected, which is present in a sample, to an antibody or antigen with which fine particles have been sensitized, and then measuring the agglutination state of the fine particles thus obtained. This type agglutination method has generally been used because of its allowing visual judgement.
There has also been employed a radioimmunoassay, an enzyme immunoassay or a fluorescent immunoassay, which comprises binding, by immunoreaction, a substance to be detected, which is present in a sample, to an antibody or antigen labelled with a labelling substance consisting of a radioactive isotope, an enzyme or a fluorescent substance, respectively, and detecting the labelling substance in the bound state.
In these immunoassays, a competitive reaction or a sandwich reaction has been widely used. Immunochromatography is known as a so-called sandwich type immunoassay. In a typical immunochromatography, the following procedure is carried out in order to detect a substance to be detected consisting of an antigen in a sample:
(1) Fine particles sensitized with an antibody to an antigen, where the antigen is the substance to be detected, are immobilized as solid phase fine particles in a chromatographic medium or an antibody is immobilized directly on a chromatographic medium to thereby prepare a chromatographic medium having a reaction site or sites. PA1 (2) Separately, fine particles are sensitized with an antibody which is specifically bindable to the substance to be detected, to thereby prepare fine particles having a labelling substance (said fine particles are referred to hereinafter as "labelling fine particles"). PA1 (3) The sensitized labelling fine particles are allowed to move chromatographically, together with a sample, in the chromatographic medium.
In the above procedure, the antibody acts as an immobilized reagent at the reaction site or sites formed in the chromatographic medium, and the sensitized labelling fine particles are specifically bound to the immobilized reagent via an antigen, which is the substance to be detected. As a result, the sensitized labelling fine particles are captured at the reaction site or sites, and the generation of a signal thereby or the intensity of the signal generated is judged by the naked eye, whereby the presence or absence and amount of the substance to be detected in the sample are determined.
In such an immunochromatography, as the fine particles used for the preparation of labelling fine particles, there have been used colloidal metal or metal oxide particles of gold, platinum, copper, iron oxide or the like; colloidal non-metal particles of sulfur or the like; and dye particles.
However, when the colloidal metal or metal oxide particles or colloidal non-metal particles are used as labelling fine particles, it is impossible to obtain labelling fine particles having a desired vivid and deep color tone because the color tone is determined by the conditions for preparing said colloidal metal or metal oxide particles or colloidal non-metal particles and the particle diameters of the colloidal metal or metal oxide particles or colloidal non-metal particles.
Meanwhile, when the dye particles are used as labelling fine particles, the color tone and depth can be selected as desired. However, their dispersion stability in water is low and difficult to control, so that the sensitization of the dye particles with an antibody is not easy. In addition, even if sensitized, the labelling fine particles obtained are difficult to resuspend and have insufficient stability. Hence, the labelling fine particles are difficult to move in an immunochromatographic medium. Moreover, even if resuspended, no uniform chromatographic movement can be achieved because the distribution of particle diameters is broad. In this respect, the dye particles are not desirable.
Thus, when the labelling fine particles consisting of colloidal metal or metal oxide particles, colloidal non-metal particles or dye particles are used, it is difficult to obtain a signal of desired intensity at the reaction site or sites of the immunochromatographic medium, or a non-uniform pattern of signal is formed. Accordingly, accurate visual judgement is difficult and it is impossible to obtain a high detection sensitivity.